Capillary electrophoresis (CE) is a technique of considerable interest in the analysis of biological mixtures as it provides a number of distinct advantages over other separation processes. One advantage of capillary electrophoresis is the small volume of the capillary tube interior. This permits one to perform separations on extremely small volumes, i.e., on volumes ranging anywhere from a few nanoliters of sample to the cytosolic fluid of a single cell (T. M. Olefirowicz and A. G. Ewing, Anal. Chem., 62:1872-1876 (1990)). Another advantage of capillary electrophoresis is the rapid rate at which heat is dissipated outward from the capillary tube due to the capillary's narrow bore. This permits the use of a high voltage to drive the electrophoresis which, in turn, provides for separations at high speed and with high efficiency and resolution. Each of these advantages renders capillary electrophoresis particularly useful for analyzing samples of biological interest, particularly mixtures of peptides, proteins, and nucleic acids.
Moreover, capillary isoelectric focusing (CE-IEF) is a rapid and high resolution separation technique which can resolve proteins based on small differences in isoelectric points. CE-IEF has been applied to the separation of hemoglobins (S. Hjerten and M. Zhu, J. Chromatogr., 346:265-270 (1985)), transferfins (F. Kilar and S. Hjerten, Electrophoresis, 10:23-29 (1989)), and immunoglobulins (Wehr, et al., Am. Biotech. Lab., 8:22-29 (1990)). Although useful, an obvious disadvantage with all methods based on IEF is that many proteins precipitate at their isoelectric points, particularly at high protein and salt concentrations, and at elevated temperatures. More particularly, it is known that the presence of salt in a sample changes the pH gradient and confines the protein zone into a small segment of the capillary. This narrow pH gradient results in high protein concentrations and, in turn, in an increased risk of precipitation, loss of resolution and long mobilization times (Zhu, et al., J. Chromatogr., 559:479-488 (1991)). In addition, this narrow pH gradient contributes to localized overheating and, thus, to irreproducibility.
For the foregoing reasons, desalting of biological samples prior to capillary electrophoresis and, in particular, capillary isoelectric focusing is highly recommended. Unfortunately, the currently used desalting techniques frequently result in large sample losses when the sample volume is below 5 .mu.l. To date, no desalting methods are available for sample volumes in the nanoliter range. As such, there still remains a need in the art for methods for the microscale desalting of biological samples. The present invention satisfies this need by providing such methods.